CN108531586B - Circulating miRNA marker on X chromosome related to breast cancer auxiliary diagnosis and application thereof - Google Patents
Circulating miRNA marker on X chromosome related to breast cancer auxiliary diagnosis and application thereof Download PDFInfo
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Abstract
The invention discloses a circulating miRNA marker on an X chromosome related to breast cancer auxiliary diagnosis and application thereof, the marker is one or more of plasma markers miR-106a-3p, miR-106a-5p, miR-20b-5p, miR-92a-2-5p, miR-500a-5p, miR-501-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p and serum markers miR-106a-5p, miR-19b-3p, miR-20b-5p, miR-92a-3p, miR-501-3p, miR-502-3p, miR-532-3p, miR-532-5p and miR-188-3 p. The circulating miRNA is used as a novel biomarker and has the characteristics of good stability, easy minimally invasive acquisition, and high sensitivity and specificity. The development and utilization of the molecular markers can provide a new direction for the diagnosis and further treatment of various diseases including tumors. The research can more specifically obtain the circulating miRNA markers of the breast cancer with clinical diagnosis potential. The study demonstrated the reliability and reproducibility of this group of mirnas as noninvasive markers for the diagnosis of breast cancer.
Description
Technical Field
The invention belongs to the field of genetic engineering and oncology, and relates to a circulating miRNA marker on an X chromosome, which is related to breast cancer auxiliary diagnosis, and an application thereof.
Background
Breast Cancer (BC) is the most common malignancy in women, and accounts for 25.2% of all worldwide malignancies according to the latest World Cancer Report 2014. In China, the incidence of breast cancer is on the rising trend, and in 2015, the incidence of breast cancer of Chinese women is the first of malignant tumors and the fatality rate is the sixth. The existing examination means such as breast ultrasonic examination, molybdenum target examination, nuclear magnetic resonance examination and hollow needle aspiration biopsy all need a certain volume of tumor tissue to detect, and have certain limitation in early diagnosis of breast cancer; the tumor markers commonly used in clinic, such as carbohydrate antigen 15-3(CA15-3), are not highly sensitive or specific. With the development of biotechnology such as genomics, proteomics, and metabolomics, more and more biomarkers have been discovered or studied. Therefore, the discovery of a new marker capable of early diagnosis of breast cancer is indicated as "Nissan", thereby facilitating early intervention and treatment of breast cancer and prolonging the life of patients.
Micro RNA (miRNAs) is a small non-coding RNA molecule with evolution conservation and about 18-25 nucleotides in length, and is widely involved in a plurality of physiological and pathological processes of organisms including tumor occurrence, invasion, metastasis and the like through regulation and control after transcription. The expression of mirnas is both time-specific and tissue-specific, and some mirnas can be involved in specific physiopathologies as well as in specific disease processes. Researches show that the expression of miRNA has different degrees of up-regulation and down-regulation in tumors, and lays a foundation for the miRNA to be used as a new tumor marker. In 2008, Mitchell detected free mirnas in peripheral blood, which were found to be able to stably exist in peripheral blood and could be used as a non-invasive marker for diagnosing tumors. This finding has opened the way that numerous researchers around the world have begun exploring circulating mirnas as noninvasive markers. The existing research proves the potential diagnosis value of circulating miRNA in breast cancer, gastric cancer, lung cancer, ovarian cancer, colorectal cancer and other tumors. However, the results of the study are not completely consistent due to differences in the study methods and the included population. Therefore, the research aims to search circulating miRNA with potential diagnosis value for breast cancer through the research on the serum and plasma of breast cancer with large samples based on the absolute quantitative method of qRT-PCR. And the expression of the miRNA in breast cancer tissues and peripheral exosomes is verified so as to further define the relation between the miRNA and the breast cancer. If a diagnostic kit aiming at breast cancer is designed according to the miRNA, the diagnosis and treatment level of breast cancer in China can be promoted, and a thought is provided for further research on breast cancer in the future.
Disclosure of Invention
The invention aims to provide a circulating miRNA marker located on an X chromosome and related to breast cancer auxiliary diagnosis.
The invention also aims to provide the circulating miRNA marker and the application of the primer thereof in preparing an auxiliary diagnosis kit for breast cancer and a medicament for treating breast cancer.
The invention also aims to provide a kit and a medicament for auxiliary diagnosis and treatment of breast cancer.
The purpose of the invention can be realized by the following technical scheme:
a circulating miRNA marker related to breast cancer aided diagnosis is a plasma marker miR-106a-3p (CUGCAAUGUAAGCAUUAC), miR-106a-5p (GUAAGUGCUUAGUGCAGGUAG), miR-20b-5p (CAAAGUGCUCAUAGUGCAGGUAG), miR-92a-2-5p (GGGUGGGGAUUUGCAUUAC), miR-500a-5p (UAAUCCUUGGCUACCUGGGUGGGUGAGA), miR-501-5p (GUUCUUCUGCCUGUCUGUGUGUGGGUGUGAGUGA), miR-188-3p (GGUCCACACACAGGGUUUGAGUG), miR-7 f-5p (GUAGGUAGGUAGGUAGUAGAGCAUUG) and miR-98-5p (GUAGGUAGGUAGGUAGUGAUGUAGUGUA) and a serum markers miR-106a-5p (GGGUAAGUAAGUAGGUAGGUAGGUAGGUAGGUAGUGCAAUUGCAAUUGCAUUG-3), miR-5 p (GUAAGUAGUGCAAUUGCAAUUGCAAUUGCAUGAGUG-3, miR-5 p) and GUAGUGCAAUUGCAAUUGCAAUUGCAUGAGUG-5 p (GUAGUGCAAUUGCAAUUGCAAUUGCAAUUG-3, GUAGUGCAAUUGCAAUUGCAAUUG-5) and GUAGUGAGUGAGUGAGUGAGUGAGUGCAAUUGCAAUUG-3, miR-5 AGUGCAAUUGCAAUUGCAAUUGAGUGAGUGCAAUUG-3, miR-5 p, GUAGUGCAAUUGCAAUUGCAAUUGCAAUUGCAAUUGCAAUUGCAAUUGCAAUUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGCAAUUGAGUGCAAUUGCAUUGAGUGAGUG-3 and CAAUUGAGUGAGUGAGUGAGUGAGUGAGUGAGUG-3 and CAAUUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUG-5P, GUAGUGAGUG-3 and GUAGUGAGUGCAAUUGAGUGAGUGAGUGAGUGAGUGCAAUUGCAAUUGAGUGAGUGAGUGAGUGAGUGAGUGAGUG-3 and CAAUUGAGUGAGUG-5P, and CAAUUGAGUGCAAUUGCAAUUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGCAAUCAAUCAAUCAAUCAAUCAAUCAAUCAAUCAAUCAAUUGAGUGAGUGAGUGAGUGAGUGAGUGAGUGAGUG-3 and CAAUGUAGUGAGUGAGUGAGUGAGUGAGUGAGCAAUCAAUCAAUCAAUCAAUC-3 and CAAUUGAGUGAGCAAUCAAUC and CAAUC-3 and CAAUUGAGCAAUUGAGUGAGCAAUC-3 and CAAUGGAGUGAGUGAGUGAGUGAGUGAGUG-3 and CAAUUGAGUGAGCAAUC-3 and CAAUUGAGUGAG, one or more of miR-532-3p (CCUCCCACACACCCAAAGGCUUGCA), miR-532-5p (CAUGCCUUGAGUGAGACCGU) and miR-188-3p (CUCCCACACUGCAGGGGUUUGCA). The circulating miRNA markers are preferably combinations of two or more of plasma markers miR-106a-3p, miR-106a-5p, miR-20b-5p, miR-92a-2-5p, miR-500a-5p, miR-501-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p and serum markers miR-106a-5p, miR-19b-3p, miR-20b-5p, miR-92a-3p, miR-501-3p, miR-502-3p, miR-532-3p, miR-532-5p and miR-188-3p, further preferably plasma markers miR-106a-3p, a combination of eighteen miRNAs of miR-106a-5p, miR-20b-5p, miR-92a-2-5p, miR-500a-5p, miR-501-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p and blood serum miR-106a-5p, miR-19b-3p, miR-20b-5p, miR-92a-3p, miR-501-3p, miR-502-3p, miR-532-3p, miR-532-5p and miR-188-3 p.
The circulating miRNA marker is applied to auxiliary diagnosis of breast cancer.
The circulating miRNA marker is applied to the preparation of an auxiliary breast cancer diagnosis kit or a medicine for treating breast cancer.
Primers of circulating miRNA markers related to breast cancer aided diagnosis, wherein the primers comprise primers of one or more miRNAs in plasma miR-106a-3p, miR-106a-5p, miR-20b-5p, miR-92a-2-5p, miR-500a-5p, miR-501-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p and serum miR-106a-5p, miR-19b-3p, miR-20b-5p, miR-92a-3p, miR-501-3p, miR-502-3p, miR-532-3p, miR-532-5p and miR-188-3 p; preferably primers containing two or more miRNAs in plasma miR-106a-3p, miR-106a-5p, miR-20b-5p, miR-92a-2-5p, miR-500a-5p, miR-501-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p and serum miR-106a-5p, miR-19b-3p, miR-20b-5p, miR-92a-3p, miR-501-3p, miR-502-3p, miR-532-3p, miR-532-5p and miR-188-3 p; further preferred are primers comprising eighteen miRNAs of plasma miR-106a-3p, miR-106a-5p, miR-20b-5p, miR-92a-2-5p, miR-500a-5p, miR-501-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p and serum miR-106a-5p, miR-19b-3p, miR-20b-5p, miR-92a-3p, miR-501-3p, miR-502-3p, miR-532-3p, miR-532-5p and miR-188-3 p.
The primer is applied to auxiliary diagnosis of breast cancer or preparation of an auxiliary diagnosis kit for breast cancer.
A breast cancer auxiliary diagnosis kit contains primers of one or more miRNAs in plasma miR-106a-3p, miR-106a-5p, miR-20b-5p, miR-92a-2-5p, miR-500a-5p, miR-501-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p and serum miR-106a-5p, miR-19b-3p, miR-20b-5p, miR-92a-3p, miR-501-3p, miR-502-3p, miR-532-3p, miR-532-5p and miR-188-3 p; preferably primers containing two or more miRNAs in plasma miR-106a-3p, miR-106a-5p, miR-20b-5p, miR-92a-2-5p, miR-500a-5p, miR-501-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p and serum miR-106a-5p, miR-19b-3p, miR-20b-5p, miR-92a-3p, miR-501-3p, miR-502-3p, miR-532-3p, miR-532-5p and miR-188-3 p; further preferably, the primer contains eighteen miRNAs of plasma miR-106a-3p, miR-106a-5p, miR-20b-5p, miR-92a-2-5p, miR-500a-5p, miR-501-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p and serum miR-106a-5p, miR-19b-3p, miR-20b-5p, miR-92a-3p, miR-501-3p, miR-502-3p, miR-532-3p, miR-532-5p and miR-188-3 p.
The kit also comprises reagents commonly used in PCR technology, such as reverse transcriptase, buffer solution, dNTPs, MgCl2DEPC water and Taq enzyme, etc.; standards and/or controls may also be included.
The invention relates to circulating miRNA markers miR-106a-3p, miR-106a-5p, miR-20b-5p, miR-92a-2-5p, miR-500a-5p, miR-501-5p, miR-188-3p, miR-let-7f-5p, miR-98-5p, miR-19b-3p, miR-92a-3p, miR-501-3p, miR-502-3p, miR-532-3p and miR-532-5p related to breast cancer diagnosis, the sequence of each miRNA is disclosed, however, the combination of the miRNA markers as auxiliary diagnosis markers of breast cancer requires creative efforts of the technicians in the field. Amplification primers of all miRNA markers can be obtained by market purchase, and the primers of the circulating miRNA markers used in the embodiment of the invention are specific miRNA stem-loop RT-PCR primers synthesized and produced by Sharp company, Guangzhou.
Specifically, the technical solution of the present invention to solve the problem includes: (1) establishing a unified specimen library and a database: standard procedures (SOP) were used to collect blood samples meeting the standards and the system collected complete demographic and clinical data. (2) Differential expression profiling of serum and plasma mirnas: circulating miRNAs differentially expressed in breast cancer and normal control populations were analyzed, and further large sample multi-stage validation of the differentially expressed miRNAs was performed. (3) The ability of these mirnas to diagnose breast cancer is clear through multi-stage validation. (4) The development of a circulating miRNA diagnosis kit: miRNAs diagnostic kit is developed according to the differential expression of miRNA in serum and plasma of breast cancer patients and normal people, and noninvasive auxiliary diagnosis of the breast cancer patients is realized. (4) The expression conditions of the miRNAs in breast cancer tissues and exosomes are analyzed, the relation between the miRNAs and breast cancer is disclosed, and a basis is provided for developing medicaments possibly related to the miRNAs for treating the breast cancer in the future.
The inventor collects blood samples meeting the standard by a Standard Operation Procedure (SOP), systematically collects complete demographic data and clinical data, and adopts a qRT-PCR method and the like.
The experimental method of research mainly includes the following parts:
1. study sample selection: patients who are initially treated, have not undergone surgery or radiotherapy and chemotherapy intervention and are pathologically confirmed to be breast cancer. The normal control is a normal population for physical examination in a hospital.
2. Training set and verification set: RNA extraction is carried out on each serum and plasma sample by using an AM1556 kit (ABI company), a cDNA sample is obtained through reverse transcription reaction, and a PCR primer and SYBR Green fluorescent dye are added for PCR reaction. And (5) comparing the Ct values of the standard substance to obtain the miRNA content in the sample.
3. The RNA in breast cancer and tissues beside the cancer is extracted by using a TRIZOL-LS reagent, the RNA in exosomes is extracted by using an ExoQuick kit (SBI company) and an AM1556 kit (ABI company), and the expression difference of miRNA in the tissues and the exosomes is detected by a qRT-PCR method.
4. Statistical analysis: exercise chi2Tests, paired t tests, and non-parametric rank-sum tests compare the differences in miRNA expression levels among different study groups. The diagnostic value of serum and plasma miRNA is verified by calculating ROC curve analysis.
The research group of the invention has found a group of 18 breast cancer circulating microRNA markers (plasma markers miR-106a-3p, miR-106a-5p, miR-20b-5p, miR-92a-2-5p, miR-500a-5p, miR-501-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p as well as serum markers miR-106a-5p, miR-19b-3p, miR-20b-5p, miR-92a-3p, miR-501-3p and miR-502-3 p) with clinical diagnosis potential through carrying out systematic expression analysis on miRNA in peripheral serum and plasma of a breast cancer patient, miR-532-3p, miR-532-5p and miR-188-3 p).
The invention has the beneficial effects that:
1. compared with the traditional tumor marker, the circulating miRNA is used as a novel biomarker, and has the characteristics of good stability, easy minimally invasive acquisition, high sensitivity and high specificity. The development and utilization of the molecular markers can provide a new direction for the diagnosis and further treatment of various diseases including tumors.
2. Researchers carry out rigorous and multistage verification and evaluation on differential expression miRNA in serum and plasma of breast cancer and normal control population through a qRT-PCR relative quantification method. The reliability and repeatability of this group of mirnas as noninvasive markers for diagnosing breast cancer were confirmed.
3. Researchers find that the expressions of miR-106a-5p and miR-20b-5p in breast cancer tissues and the expressions of miR-106a-5p in blood plasma and blood serum are both obviously increased; meanwhile, the expressions of miR-106a-3p, miR-106a-5p, miR-92a-2-5p, miR-500a-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p in blood plasma and miR-106a-5p, miR-19b-3p, miR-20b-5p, miR-92a-3p, miR-501-3p, miR-502-3p and miR-188-3p in breast cancer exosomes are also obviously higher than those of normal people. The close relationship between this group of mirnas and breast cancer is shown. In addition, the expression level of plasma miR-106a-3p, miR-106a-5p, miR-20b-5p and miR-92a-2-5p in stage I and II breast cancer patients is obviously higher than that of stage III breast cancer patients, the expression level of plasma miR-106a-5p, miR-20b-5p and miR-92a-2-5p in hormone receptor positive patients is far higher than that of hormone receptor negative patients, and the expression level of plasma miR-106a-5p and miR-20b-5p in HER2 non-amplification patients is far higher than that of HER2 amplification patients. These results will provide new ideas for future studies on the mechanism of these mirnas for breast cancer and for the treatment of breast cancer with these mirnas.
Drawings
FIG. 1: flow chart of experiment
FIG. 2: 9 miRNAs highly expressed in plasma samples of breast cancer patients
FIG. 3: 9 miRNAs highly expressed in serum sample of breast cancer patient
FIG. 4: ROC curve analysis of the obtained plasma miRNA of breast cancer patients
Wherein, A1: training a set; a2: a verification set; a3: collection of training set and verification set
FIG. 5: ROC curve analysis of the obtained serum miRNA of breast cancer patients
Wherein, B1: training a set; b2: a verification set; b3: collection of training set and verification set
FIG. 6: differential expression of plasma miRNAs in different histological stages, hormone receptor status and the presence or absence of amplification of HER2 in breast cancer patients
Wherein, A: histology staging; b: a hormone receptor state; c: HER2 presence or absence of amplification status
FIG. 7: miR-106a-5p and miR-20b-5p are highly expressed in breast cancer tissues
FIG. 8: expression of 9 miRNAs in plasma samples in exosomes of breast cancer patients
FIG. 9: expression of 9 miRNAs in serum sample in exosomes of breast cancer patients
Detailed Description
The inventor collects a large amount of venous serum and plasma samples of breast cancer patients and normal physical examination people from the first subsidiary hospital of Nanjing medical university in 2014 to 2016, and selects 290 samples of breast cancer and 402 samples of normal control as qRT-PCR verified experimental samples from the samples by collating sample data. At the same time, 32 breast cancer tissues and 32 tissues beside the cancer are reserved. Selected patient serum samples were all from patients who were initially treated, were not operated, and had undergone chemo-and-radiotherapy intervention and were pathologically confirmed to be breast cancer. And the system collects the demographic data and clinical data of the samples.
Referring to the flow chart (fig. 1), 32 mirnas located on the X chromosome from three clusters are verified by a relative quantification method based on qRT-PCR through a training set and a verification set, and the specific steps are as follows:
1. serum/plasma RNA extraction: serum RNA extraction kit (AM1556) of ABI company is selected, 200ul of RNA is extracted from each sample according to the kit instruction, and finally 100ul of DEPC water is used for dissolving.
Preparation of cDNA:
1) reverse transcription experiment was performed using a 50. mu.L reaction system
The above reaction system was mixed well and after instantaneous centrifugation, the reaction was carried out according to the following procedure:
2) the following reactants are added into the reaction system after the reaction
3.qPCR
1) Using a 5. mu.L reaction system, the following ratio was used for the test
The reaction system is mixed evenly, placed in a real-time quantitative PCR instrument after instantaneous centrifugation, and reacted according to the following procedures:
the dissolution profile was added after the reaction was complete.
And (3) data analysis: statistical analysis was performed using SPSS 16.0 software to obtain a set of 9 plasma mirnas that were consistently highly expressed in breast cancer circulation in both training and validation sets: miR-106a-3p, miR-106a-5p, miR-20b-5p, miR-92a-2-5p, miR-500a-5p, miR-501-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p and 9 sera: miR-106a-5P, miR-19b-3P, miR-20b-5P, miR-92a-3P, miR-501-3P, miR-502-3P, miR-532-3P, miR-532-5P and miR-188-3P (P value is less than 0.05 in training set and validation set, and is shown in figure 2 and figure 3). From these 18 mirnas, ROC curves for each sample can be calculated. As shown in fig. 4 and fig. 5, the molecular markers composed of the 18 mirnas can well distinguish breast cancer patients from normal population. And through subgroup analysis, the expression level of plasma miR-106a-3p, miR-106a-5p, miR-20b-5p and miR-92a-2-5p in stage I and II breast cancer patients is obviously higher than that of stage III breast cancer patients, the expression level of plasma miR-106a-5p, miR-20b-5p and miR-92a-2-5p in hormone receptor positive patients is far higher than that of hormone receptor negative patients, and the expression level of plasma miR-106a-5p and miR-20b-5p in HER2 non-amplified patients is far higher than that of HER2 amplified patients (FIG. 6).
The expression of the 18 miRNAs in exosomes in breast cancer tissues, serum and plasma is further detected after the research group, TRIZOL is used for extracting RNA from the breast cancer tissues, and the exosome extraction kit is an ExoQuick kit (SBI company). Exosome RNA was extracted using AM1556 kit (ABI) after 200ul of exosome extracted from serum/plasma was resuspended in 200ul of DEPC water, in the same procedure as the serum/plasma RNA extraction.
The expression of miR-106a-5p and miR-20b-5p in breast cancer tissues is higher than that in paracarcinoma tissues by using nonparametric test analysis (figure 7). The expression of miR-106a-3p, miR-106a-5p, miR-92a-2-5p, miR-500a-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p in plasma and miR-106a-5p, miR-19b-3p, miR-20b-5p, miR-92a-3p, miR-501-3p, miR-502-3p and miR-188-3p in serum in breast cancer exosomes is also obviously higher than that in normal population (figure 8 and figure 9).
The kit comprises a batch of serum and plasma miRNA qRT-PCR primers, and can also comprise common reagents required by corresponding PCR technologies, such as: reverse transcriptase, buffer, dNTPs, MgCl2, DEPC water, fluorescent probes, RNase inhibitors, Taq enzyme and the like can be selected according to the specific experimental method, the common reagents are well known to those skilled in the art, and in addition, standard substances and controls (such as quantitative standard normal human samples and the like) can be provided. The kit has the value that only serum/plasma is needed without other tissue samples, and the expression content of miRNA in the serum/plasma sample is detected by the simplest fluorescence method, so as to assist in diagnosing the possibility of breast cancer of a patient from which the sample is derived. The miRNA detection of serum and plasma is convenient, the quantification is accurate, and the sensitivity and the specificity of disease diagnosis are greatly improved, so that the kit can help to guide diagnosis and further individualized treatment when put into practice.
Claims (2)
1. The application of a primer for detecting a circulating miRNA marker on an X chromosome in the preparation of a breast cancer auxiliary diagnosis kit is characterized in that the circulating miRNA marker is prepared from plasma markers miR-106a-3p, miR-106a-5p, miR-20b-5p, miR-92a-2-5p, miR-500a-5p, miR-501-5p, miR-188-3p, miR-let-7f-5p and miR-98-5p, serum markers miR-106a-5p, miR-19b-3p, miR-20b-5p, miR-92a-3p, miR-501-3p, miR-502-3p and miR-532-3p, miR-532-5p and miR-188-3 p.
2. The use of claim 1, wherein the kit further comprises reagents commonly used in PCR technology.
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| CN101921760A (en) * | 2010-09-08 | 2010-12-22 | 南京医科大学 | Serum/plasma miRNA marker associated with breast cancer and application thereof |
| CN105308189A (en) * | 2013-04-15 | 2016-02-03 | 瑞泽恩制药公司 | Markers of tumor cell response to anti-cancer therapy |
| CN107429295A (en) * | 2015-03-09 | 2017-12-01 | 新加坡科技研究局 | The method of risk to suffer from breast cancer is determined by detecting the expression of Microrna (miRNA) |
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| CN105308189A (en) * | 2013-04-15 | 2016-02-03 | 瑞泽恩制药公司 | Markers of tumor cell response to anti-cancer therapy |
| CN107429295A (en) * | 2015-03-09 | 2017-12-01 | 新加坡科技研究局 | The method of risk to suffer from breast cancer is determined by detecting the expression of Microrna (miRNA) |
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